Discovery of low-affinity preproinsulin epitopes and detection of autoreactive CD8 T-cells using combinatorial MHC multimers.
J Autoimmun
; 37(3): 151-9, 2011 Nov.
Article
em En
| MEDLINE
| ID: mdl-21636247
ABSTRACT
Autoreactive cytotoxic CD8 T-cells (CTLs) play a key pathogenic role in the destruction of insulin-producing beta-cells resulting in type 1 diabetes. However, knowledge regarding their targets is limited, restricting the ability to monitor the course of the disease and immune interventions. In a multi-step discovery process to identify novel CTL epitopes in human preproinsulin (PPI), PPI was digested with purified human proteasomes, and resulting COOH-fragments aligned with algorithm-predicted HLA-binding peptides to yield nine potential HLA-A1, -A2, -A3 or -B7-restricted candidates. An UV-exchange method allowed the generation of a repertoire of multimers including low-affinity HLA-binding peptides. These were labeled with quantum dot-fluorochromes and encoded in a combinatorial fashion, allowing parallel and sensitive detection of specific, low-avidity T-cells. Significantly increased frequencies of T-cells against four novel PPI epitopes (PPI(4-13)/B7, PPI(29-38)/A2, PPI(76-84)/A3 and PPI(79-88)/A3) were detected in stored blood of patients with recent onset diabetes but not in controls. Changes in frequencies of circulating CD8 T-cells against these novel epitopes were detected in blood of islet graft recipients at different time points after transplantation, which correlated with clinical outcome. In conclusion, our novel strategy involving a sensitive multiplex detection technology and requiring minimal volumes of stored blood represents a major improvement in the direct ex-vivo characterization and enumeration of immune cells in the pathogenesis of type 1 diabetes.
Texto completo:
1
Base de dados:
MEDLINE
Assunto principal:
Peptídeos
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Precursores de Proteínas
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Autoimunidade
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Linfócitos T CD8-Positivos
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Técnicas de Química Combinatória
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Diabetes Mellitus Tipo 1
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Células Secretoras de Insulina
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Insulina
Tipo de estudo:
Clinical_trials
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Diagnostic_studies
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Prognostic_studies
Idioma:
En
Revista:
J Autoimmun
Assunto da revista:
ALERGIA E IMUNOLOGIA
Ano de publicação:
2011
Tipo de documento:
Article
País de afiliação:
Holanda